Therapeutic administration of soluble CD4 is theoretically an attractive means of competitively inhibiting the binding of HIV to its primary cellular receptor, but was a failure in clinical trials more than a decade ago. We propose the creation of novel CD4 analogs to overcome the stoichiometric and affinity limitations of that initial approach. Using electrophilic phosphonate ester probes, we recently discovered that HIV gp120 contains activated nucleophilic amino acids. In the proposed studies, CD4 (and CD4 peptide mimetic, s) will be modified by the addition of activated electrophilic phosphonates, which will result in specific covalent bonding of these constructs to nucleophilic sites on gp120 on the surface of HIV virions. Such an "infinite" affinity interaction will result in irreversible blocking of HIV infectivity. The concept is based on our successful analogous work on catalytic antibodies, in which nucleophilic immunoglobulin sites bind to electrophiles in the substrate ligand. Thus the Specific Aims of this proposed Innovation Grant are (1) to create covalently reactive CD4 and CD4 peptide mimetics; (2) to determine the potency, intra- and intersubtype breadth and irreversibility of HIV neutralization and inhibition of cell-to-cell spread of HIV infection via cell fusion by these covalently reactive CD4 analogues; (3) to demonstrate the irreversibility of binding of the constructs to gp120 and to intact virions; and (4) to determine the role in the virus neutralization that isplayed by the covalency (vs. conventional noncovalent antibody binding). Successful results should lead tofollow-up testing in primate models and human clinical trials. The proposed studies may also help to definecovalent and pseudocovalent forces as a novel mechanism of protein-ligand interactions. [unreadable] [unreadable]